OBJECTIVE: To explore the therapeutic effect and target of atractylenolide I (AT-I) on post-infectious irritable bowel syndrome (PI-IBS) rats. METHODS: Therefore, the preliminarily mechanism of AT-I in anti-PI-IBS were first predicted by network pharmacology and molecular docking, then the possible signaling pathways were systematically analyzed. Finally, the potential therapeutic targets and possible signaling pathways of AT-I on PI-IBS in Sprague-Dawley (SD) rat model were verified by experiments. RESULTS: AT-I could alleviate PI-IBS symptoms and reduce the expression of tumor necrosis factor α, interleukin-6 and Interferon-gamma in PI-IBS SD rat model and inhibit the c-Jun N-terminal kinase/inducible nitric oxide synthase (JNK/iNOS) pathway. Notably, AT-I treatment could inhibit the overexpression of polymerase I and transcript release factor (PTRF). CONCLUSION: AT-I could alleviate PI-IBS symptoms through downregulation of PTRF and inhibiting the JNK/ iNOS pathway. This study not only provides a scientific basis to clarify the anti-PI-IBS effect of AT-I and its mechanism but also suggests a novel promising therapeutic strategy to treat the PI-IBS.